Thyroid hormones (TH), 3,5,3′-triiodothyronine (T3) and thyroxine (T4), are essential for growth, development and metabolism. A common way to study TH action is through the use of a wide range of TH-responsive cell lines from mammalian and other vertebrate species (e.g. rat GH3 cells and frog XTC-2 and XLA cells). Typically, these cell lines are maintained in bovine or fetal calf serum which contains sufficient levels of TH to require special treatment (“stripping”) of the serum to remove it before TH-dependent effects can be studied.
The stripping method typically requires the use of either activated charcoal or AG1-X10 or AG1-X8 resin (e.g. (Samuels, Stanley et al. 1979)). These methods are proposed for example by Lewis and Parsons U.S. Pat. No. 4,431,741 issued Feb. 14, 1984, Eisentraut U.S. Pat. No. 3,776,698 issued Dec. 4, 1973, Turner et al. U.S. Pat. No. 3,922,145 issued Nov. 25, 1975, and Hollander U.S. Pat. No. 3,928,553 issued Dec. 23, 1975. Unfortunately, these methods are not specific for TH and result in the removal of many other endogenous serum proteins, growth factors and hormones. Alteration of the serum growth medium in this way can influence the growth and survival characteristics of the cells, typically to a detrimental effect. This is exemplified by the fact that TH-responsive cells need to be maintained in regular (TH-containing) serum until a day or two before a TH-induction experiment. The growth medium is then changed to medium containing stripped serum, the cells acclimated to a reduced TH-environment and then TH is added exogenously. Incubation of cells in the stripped medium cannot be accomplished over the long term, because of the lack of necessary growth factors and hormones in the growth medium. The varied removal of these compounds also confounds interpretation and applicability to intact organisms since they themselves influence TH action (Yen 2001).
Production of hypothyroid serum is not easily achieved in mammals, since this represents a serious disease state (Yen 2001). However, there are several other vertebrates that have developmental stages wherein they are naturally in a functionally athyroid state while in a growth intensive phase of their normal development (Norris 1997). Some examples of this occur in frog tadpoles (Regard, Taurog et al. 1978; Kaltenbach 1996), salmonids prior to smoltification (Dickhoff and Sullivan 1987; Specker 1988; Eales and Brown 1993), flatfish (Inui and Miwa 1985), salamanders (Safi, Begue et al. 1997), and lampreys (Youson and Sower 2001).
To illustrate the point further in Amphibia, the larval phase of postembryonic development is primarily a period of extensive growth in the absence of a functional thyroid gland. This premetamorphic phase is followed by a prometamorphic phase in which the thyroid gland matures and low-level secretion of TH occurs (White and Nicoll 1981). TH levels rise and peak at metamorphic climax, which is characterized by the rapid, overt remodeling of the tadpole. It is these growth-intensive periods of development which are conducive to the production of serum with naturally low levels of THs. Low levels of thyroid hormone are considered to be less than approximately 100 ng/dl total T3 or 5 micrograms/dl total T4 as these are the lower end of normal range for humans and cattle (Samuels, Stanley et al. 1979; Shanker, Rao et al. 1984; Health 2005).
Despite the foregoing, there have been no attempts to produce or use serum derived from vertebrates during their athyroid, or low thyroid state. It is an object of the present embodiment to overcome the deficiencies in the prior art.